Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer mortality with near 100% of the victims succumbing within 6 month of diagnosis. New strategies for PDAC prevention are therefore urgently needed. Using two risk factors for PDAC (pancreatitis and smoking), we have established a hamster model of PDAC by inducing pancreatitis in the animals via ethanol in the drinking water while additionally injecting them with the nicotine-derived carcinogenic nitrosamine 4-(methylnitrosamino)-1-(3-pyridyl)-1- butanone (NNK). Using this animal model as well as xenografts from human PDAC in mice and in vitro studies with human PDAC cell lines and the putative cell of origin of PDAC, pancreatic duct epithelial cells, our published and new preliminary data show that adenylyl cyclase-dependent intracellular signaling downstream of beta-adrenoreceptors (b-ARs) stimulates PDAC while additionally triggering the release of epidermal growth factor, vascular endothelial growth factor and arachidonic acid. NNK activates this signaling pathway directly by binding to b-ARs and indirectly by stimulating the a7nicotinic acetylcholine receptor (a7nAChR)-mediated release of the stress neurotransmitters noradrenaline and adrenaline, which are agonists for b-ARs. All components of this stimulatory network are upregulated in PDAC while at the same time g-aminobutyric acid (GABA) which inhibits this pathway by blocking the activation of adenylyl cyclase is suppressed. Treatment of PDAC cells in vitro or PDAC xenografts in vivo with GABA had inhibiting effects. These findings suggest GABA as a potential PDAC preventive agent. To test this hypothesis and at the same time further our understanding on the regulation of PDAC by stimulatory b-AR signaling and inhibitory GABA signaling we propose four specific aims. Specific Aim 1: Using our hamster model, we will test the hypothesis that GABA or the synthetic GABA analogue baclophen prevent the development of PDAC. Specific Aim 2: Using PCR microarrays, Western blotting and immunohistochemistry, we will investigate the modulation of markers for proliferation, angiogenesis, metastasis, apoptosis, cell renewal, and cAMP signaling by GABA in tissues from hamster PDAC and normal pancreatic tissue. Specific Aim 3: Using human PDAC cell lines and pancreatic duct epithelial cells in vitro, we will determine the inhibitory actions of GABA in b-AR and PGE2-mediated signaling pathways and explore a potential reduction in TNFa and IL-1b by GABA. Specific Aim 4: We will test the hypothesis that inhibited GABA production due to NNK-induced desensitization of the a4nAChR and stimulation of stress neurotransmitter production due to NNK-induced upregulation of the a7nAChR contribute to the stimulation of PDAC. Data generated will provide a preclinical basis for the use of GABA in the prevention of PDAC. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is the fourth leading cause of cancer death with a mortality near 100% within one year of diagnosis because it does not respond to existing therapies and metastasizes extensively. New strategies to combat this deadly disease are thus urgently needed. Data presented in this project along with known biological effects of risk factors for pancreatic cancer (smoking, diabetes, pancreatitis) suggest that hyperactive beta-adrenoreceptors and possibly other G-protein coupled receptors stimulate pancreatic cancer development and progression while at the same time the pancreatic production of gamma-aminobutyric acid (GABA), which normally controls the activity of these receptors via activation of the inhibitory GABAB receptor, is reduced. Based on this novel concept, we propose to restore deficient pancreatic GABA for the prevention of pancreatic cancer. In vitro and in vivo studies proposed under this project will provide a preclinical basis for the use of GABA-ergic agents for the marker-guided prevention of pancreatic cancer in individuals with pancreatic GABA deficiency.